The invention relates generally to a tissue regeneration system, and more particularly to a system that includes a layered plurality of mineral matrices, where at least two of the layers include a biomolecule having a cell-affecting portion and a matrix-binding portion, and where the biomolecule is releasably associated with the matrix. In use, the plurality of mineral matrices degrade at various predictable rates, facilitating temporal control over release of the biomolecule(s) from the matrices.
One area of tissue regeneration that would benefit from improved biological surrogates is bone tissue regeneration systems. Under physiological conditions, bone tissue regeneration involves a complex interplay of multiple biologically active molecules and stem cells. The biologically active molecules are often presented sequentially in “cascades,” where each factor has a distinct effect on the cells of a growing bone issue. These biologically active molecules can be exploited to direct active regeneration of functional bone tissue for repair or for replacement. A key issue in designing systems to aid in bone tissue regeneration is to temporally control tissue concentration of biologically active molecules such as growth factors and/or cytokines.
Regenerating natural bone tissue represents a promising approach to bone replacement and could supplant many of the current, metallic, hardware-based bone replacement methods and expand the range of orthopedic conditions that can be effectively treated. Potential applications of novel bone regeneration systems include filling of bone voids in non-union fractures or maxillofacial deformities, bridging of gaps in spine fusion surgeries and stabilizing vertebral compression fractures. Not only would improved bone tissue regeneration systems offer an expanded range of treatment for orthopedic conditions, they would also be economically advantageous.
Existing passive bone tissue repairing or replacing systems do not exert a high level of control over the process of new bone formation. Such passive tissue regeneration systems include simply adding growth factors to a defect site in solution. However, such systems are inefficient because single growth factors delivered either by bolus injections into the site of disease or by systemic administration require very high levels for a measurable in vivo effect. In many instances, the growth factors will simply diffuse away from a defect site, leading to limited effects. Additionally, uncontrolled growth factor activity may occur at a distant site. See Yancopoulos G, et al., “Vascular-specific growth factors and blood vessel formation,” Nature 407:242-248 (2000).
To solve these problems, recent tissue regeneration systems embed growth factors into plastic microspheres, thereby localizing growth factors to a defect site. See Langer R & Moses M, “Biocompatible controlled release polymers for delivery of polypeptides and growth factors,” J. Cell Biochem. 45:340-345 (1991); Langer R, “New methods of drug delivery,” Science 249:1527-1533 (1990); Leong K, et al., “Polyanhydrides for controlled release of bioactive agents,” Biomaterials 7:364-371 (1986); Cohen S, et al., “Controlled delivery systems for proteins based on poly(lactic/glycolic acid) microspheres,” Pharm. Res. 8:713-720 (1991); and Pekarek K, et al., “Double-walled polymer microspheres for controlled drug release,” Nature 367:258-260 (1994). None of these systems, however, provides a structural matrix for tissue ingrowth. In addition, these systems are difficult to process into structural matrices while retaining adequate biological activity of the growth factor. Furthermore, many of these systems have failed to demonstrate the ability to temporally deliver multiple growth factors.
Other tissue regeneration systems embed growth factors in hydrated gels, thereby localizing growth factors to a defect site. See Lee K, et al., “Controlled growth factor release from synthetic extracellular matrices,” Nature 408:998-1000 (2000); Tabata Y & Ikada Y, “Vascularization effect of basic fibroblast growth factor released from gelatin hydrogels with different biodegradabilities,” Biomaterials 20:2169-2175 (1999); and Anseth K, et al., “In situ forming degradable networks and their application in tissue engineering and drug delivery,” J. Control. Release 78:199-209 (2002). However, like plastic microspheres, hydrated gels are not particularly well-suited for certain types of tissue regeneration because the growth factors rapidly diffuse out of the gel matrix, resulting in limited signaling.
To overcome these problems, the most recent tissue regeneration systems have relied upon methods of gas foaming a porous plastic scaffold to allow for incorporation of growth factors with biological activity and variable release rates of several days to months. See Murphy W, et al., “Sustained release of vascular endothelial growth factor from mineralized poly(lactide-co-glycide) scaffolds for tissue engineering,” Biomaterials 21:2521-2527 (2000); Murphy W, et al., “Bone regeneration via a mineral substrate and induced angiogenesis,” J. Dent. Res. 83:204-210 (2004); Sheridan M, et al., “Bioabsorbable polymer scaffolds for tissue engineering capable of sustained growth factor delivery,” J. Control. Release 64:94-102 (2000); Howdle S, et al., “Supercritical fluid mixing: preparation of thermally sensitive polymer composites containing bioactive materials,” Chemical Commun. 1:1-2 (2001); and Yang X, et al., “Novel osteoinductive biomimetic scaffolds stimulate human osteoprogenitor activity—implications for skeletal repair,” Connect. Tissue Res. 44:312-317 (2003). See also U.S. Pat. No. 6,676,928.
Similarly, others have used covalent conjugation of growth factors to hydrogels and multilayered hydrogels to provide enhanced control over osteogenic growth factor delivery. See Zisch A, et al., “Covalently conjugated VEGF-fibrin matrices for endothelialization,” J. Control. Release 72:101-113 (2001); Raiche A & Puleo D, “Cell responses to BMP-2 and IGF-I released with different time-dependent profiles,” J. Biomed. Mater. Res. 69A:342-350 (2004); and Raiche A & Puleo D, “In vitro effects of combined and sequential delivery of two bone growth factors,” Biomaterials 25:677-685 (2004). These tissue regeneration systems, however, have not yet achieved satisfactory temporal control over cell activity while new tissue forms. In addition, these tissue regeneration systems have difficultly in temporally controlling the processing of heterogeneous and degradable materials with layers containing growth factors. Furthermore, none of these systems permits an adequate release of growth factors from a single matrix using release mechanisms that occur over distinct timeframes.
For the foregoing reasons, there is a need for a tissue regeneration system that localizes and temporally controls the release of multiple growth factors to stimulate tissue regeneration.